Determination of the mechanism of biological N2 fixation will be approached in the following ways. By kinetic analysis and by using ba thophenathroline sulfonate as a probe of Fe exposed in the Fe protein of nitrogenase by added MgATP, the function of MgATP in N2 fixation will be examined. Determine the rate-limiting step and the point of MgATP hydrolysis among the electron transfer steps in N2 fixation. Examine the interactions of nitrogenase substrates and inhibitors to ascertain how they affect each other at the electron sink of nitrogenase. Study why H ion reduction by nitrogenase is not inhibited by CO, whereas the reduction of all other substrates is inhibited, and determine why C2H2 inhibits N2 reduction noncompetitively whereas N2 inhibits C2H2 reduction competitively. Seek unequivocal evidence that nitrogenase substrates all bind to the MoFe protein. Determine whether or not the "exchange" reaction between H2O and D2 supports the concept that hydrazine and diazene are intermediates in N2 reduction, and establish the influence of N2 at low pN2 on the "exchange" reaction. Study the tight binding between the MoFe protein of Azotobacter vinelandii and the Fe protein of Clostridium pasteurianum to determine the binding ratio between components and to find whether nitrogenase acts as a complex or whether it dissociates into the MoFe and Fe proteins at each turn of the cycle. Purify the activating factor for the Fe protein from Rhodospirillum rubrum and determine how it works. Determine how radiant energy is captured by the blue-green algae so that it can be used to drive biological N2 fixation.